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United States Patent |
5,160,627
|
Cussler
,   et al.
|
November 3, 1992
|
Process for making microporous membranes having gel-filled pores, and
separations methods using such membranes
Abstract
A process is provided for modifying the properties of a hydrophobic
microporous membrane which includes the steps of first providing a
hydrophobic microporous membrane, treating it with a surfactant to render
the membrane hydrophilic, wetting the membrane with an aqueous solution of
a polyol such as polyvinyl alcohol (PVA) and divinyl sulfone (DVS) or a
precursor thereof, washing the membrane with water to displace the
polyol/DVA from the exterior of the membrane while retaining it in the
pores of the membrane, and crosslinking the polyol/DVS into an aqueous gel
to yield a hydrophilic microporous membrane having pores filled with an
aqueous polyol/DVS gel, the exterior of the membrane being unobstructed by
gel. The modified membranes produced according to the process are useful
in carrying out chromatographic separations.
Inventors:
|
Cussler; Edward L. (Edina, MN);
Gillberg-LaForce; Gunilla E. (Summit, NJ);
Sansone; Michael J. (Berkeley Heights, NJ);
Schisla; David K. (St. Louis, MO)
|
Assignee:
|
Hoechst Celanese Corporation (Somerville, NJ)
|
Appl. No.:
|
815297 |
Filed:
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December 27, 1991 |
Current U.S. Class: |
210/639; 210/198.2; 210/635; 210/656; 264/DIG.48; 264/DIG.62 |
Intern'l Class: |
E01D 067/00 |
Field of Search: |
264/41,45.1,48,DIG. 48,DIG. 62
210/639
|
References Cited
U.S. Patent Documents
3335545 | Aug., 1967 | Robb et al.
| |
4113912 | Sep., 1978 | Okita.
| |
4405688 | Sep., 1983 | Lowery et al.
| |
4451981 | Jun., 1984 | Kaniarz.
| |
4541981 | Jun., 1984 | Lowery.
| |
4789468 | Dec., 1988 | Sirkar.
| |
4957620 | Sep., 1990 | Cussler.
| |
4994189 | Feb., 1991 | Leighton et al. | 210/637.
|
Foreign Patent Documents |
69869 | Jul., 1981 | EP.
| |
257635 | Aug., 1987 | EP.
| |
302650 | Jul., 1988 | EP.
| |
8801183 | May., 1988 | NL.
| |
Other References
Schisla, D., "Hollow Fiber Liquid Chromatography With A Gel Stationary
Phase", Center for Interfacial Engineering, Fall Review of the Polymer
Microstructure Program, Oct. 16-17, 1989.
Handbook of Fiber Science and Technology: vol. II-Chemical Process of
Fibers and Fabrics, Functional Finishes Part A, pp. 23-28, Marcel Dekker
Inc., NY 1983.
Ding, H., Yang, M. Schisla, D. and Cussler, E., "Hollow Fiber Liquid
Chromatography", AIChE Journal, 35 (5), May 1989, pp. 814-882.
Neplenbroek, A. M. et al, "Stable Supported Liquid Membranes", Proceedings,
vol. I, The 1990 International Congress on Membranes and Membrane
Processes, (North American Membrane Society), pp. 686-688.
|
Primary Examiner: Sever; Frank
Attorney, Agent or Firm: Genoni; K. A., Brown; J. M., Davidson; B. H.
Parent Case Text
This is a continuation of application Ser. No. 07/727,330 filed Jul. 3,
1991, now abandoned which was a continuation of Ser. No. 07/599,494, filed
Oct. 17, 1990, now abandoned.
High performance liquid chromatography is a well known method of separating
solute species in dependence upon the differential absorption/desorption
between two different solute species. Typically, a liquid carrier (in
which the solute species to be separated are present), is passed through a
column packed with separation media (e.g., solid or gelled particles).
This separation medium, in effect, increases the residence time of one (or
more) solute species in the liquid carrier (which is inert to the solute
species) relative to one (or more) other solute species in the liquid
carrier (i.e., due to the greater rate of absorption/desorption of the one
solute species relative to the other solute species). Due to the increased
residence time of the one solute species in the column, there will be a
time when an essentially pure mixture of carrier liquid and the other
solute species will be present at the discharge of the column--that is,
the one and the other solute species will be separated.
With the recent advent of commercial manufacture of biological species
(e.g. proteins), however, the conventional liquid chromatography technique
of using packed particle beds has proven to be an inefficient means of
separating one species from another with high purity. This inability of
packed particle chromatography columns can be attributed generally to the
high pressure drops which are experienced and which lead to lesser flow
rates of the liquid carrier through the column (thereby leading to a
lesser rate of production of the desired biological species). In addition,
the requirements for very stringent control over particle size, the
uniformity of the particles and the manner in which such particles fill
the column contribute to increased costs which, in general, cannot be
tolerated on a commercial scale. Furthermore, particle beds sometimes
become plugged or fouled after a number of sample injections. Hence, the
conventional packed particle technique for liquid chromatography, while
being adequate for analytical purposes on a small scale, is inadequate for
separating solute species (particularly biological species) on a
commercial scale.
Recently, it has been proposed to employ hollow fibers for liquid
chromatography. The geometry of such hollow fibers provides an attractive
alternative to particles in terms of lesser pressure drop through the
column, and the lower cost of hollow fibers ( relative to uniform
particles) which provides attractive economies of scale and thus may allow
commercially viable liquid chromatography to be achieved. Larger bed
volumes can be more easily achieved in the hollow fiber geometry. Adding
length to the column does not increase pressure drops with the severity
encountered with particle beds--a five meter module has a pressure drop
below one pound per square inch. Adding fibers in parallel gives an
efficient mobile phase and sample distribution if the fiber diameters are
well matched. The hollow fibers allow the advantage of using a low
pressure pump, and the wide open conduits of the fibers should plug less
easily than particle packing The low resistance to flow in the fibers will
allow for more speedy washing, elution, and regeneration steps for
affinity chromatography.
The art has developed in recent years concerning modification of the hollow
fiber lumens to improve and expand on the performance of hollow fibers in
chromatographic separations.
European Published Patent Application 302,650 discloses a method for
converting a hydrophobic polyolefin hollow fiber microporous membrane to a
hydrophilic membrane by the grafting of polyvinyl alcohol onto the inner
and outer surfaces of a hollow fiber substrate membrane. The method
consists of irradiation of the hollow fibers with ionizing radiation,
followed by reaction with vinyl acetate and then hydrolysis.
Okita U.S. Pat. No. 4,113,912 teaches that a fluorocarbon microporous
membrane, such as polyvinylidene fluoride or polytetrafluoroethylene, can
be made hydrophilic by filling the pores with an aqueous solution of a
water-soluble polymer, as for example polyacrylic acid, polyacrylamide, or
polyvinyl alcohol, and then subjecting the polymer-treated membrane to
reagents and conditions that lead to water-insolubilization of the
polymer, generally by crosslinking. The resulting membrane is suitable for
use in filtration, dialysis, ultrafiltration, and reverse osmosis.
European Published Patent Application 257,635 teaches that hydrophobic
membranes, with fluorocarbon membranes used as examples, can be rendered
hydrophilic by filling the pores with an aqueous solution containing one
or more hydrophilic polyfunctional amine- or hydroxy-containing monomers
or polymers, such as water-soluble cellulose derivatives or polyvinyl
alcohol, along with crosslinking agents and optional catalysts,
surfactants and initiators. These solutions are formulated with the goals
of improving penetration of the pores and also of inducing crosslinking to
take place or causing the hydrophilic compound to chemically bind as an
insoluble coating on the fluorocarbon substrate The product membranes are
useful in ultra- and microfiltration.
SUMMARY OF THE INVENTION
This invention relates to a process for modifying the properties of a
hydrophobic microporous membrane which includes the steps of first
providing a hydrophobic microporous membrane, treating it with a
surfactant to render the membrane hydrophilic, wetting the lumens of the
fibers with an aqueous solution of polyvinyl alcohol (PVA) and divinyl
sulfone (DVS), washing the lumens with water to displace the PVA/DVS from
the lumens while retaining it in the pores, injecting a base to initiate
and catalyze the crosslinking of the PVA/DVS into an aqueous gel, and
washing out the base, to yield a hydrophilic microporous membrane having
pores filled with an aqueous PVA/DVS gel, and having unobstructed lumens.
The advantage of providing an aqueous gel in the membrane pores is that the
gelled water can withstand higher pressure gradients without being
displaced from the pore structure, compared with the ungelled liquid. The
gelling of the water has only minor influence on the diffusion rates of
small molecules and/or carriers in the liquid. The liquid (water) becomes
the transport medium when the membrane is used for separations. A carrier
can also be dissolved in the immobilized water to facilitate transport of
molecules being separated in the liquid medium.
The pore-filled membranes may be used broadly in liquid chromatography
applications, especially where it is advantageous to employ a hollow fiber
membrane which can operate under high pressure drops without bleeding of
the gel out of the fiber matrix and where unimpeded diffusion of a liquid
mobile phase through the fiber lumens is important. The gel in the pores
of the hollow fibers of the invention can be used as a backbone for
affinity ligands that selectively bind with proteins, or as a selective
hydrophilic environment for separating low molecular weight biological
molecules.
It is an object of this invention to provide a stabilized gelled liquid
membrane immobilized in the pores of a microporous membrane, which can be
used in separations at high pressure without loss of the liquid membrane
by rejection from the microporous membrane matrix.
Claims
We claim:
1. A method of modifying a hydrophobic microporous membrane, by providing a
stabilized gelled liquid membrane immobilized in the pores of a
microporous membrane having properties for enabling separations at high
pressure drops across the membrane while substantially precluding loss of
the liquid membrane by rejection from the microporous matrix, comprising
the steps of:
(a) treating the membrane with a surfactant;
(b) draining excess surfactant from the membrane;
(c) drying the membrane;
(d) preparing an aqueous solution of at least one polyol and at least one
compound selected from the group consisting of divinyl sulfone and divinyl
sulfone precursors;
(e) treating the membrane with the aqueous solution;
(f) removing aqueous solution from the exterior of the membrane by washing
the membrane with water;
(g) crosslinking the aqueous solution in the pores of the membrane; and
(h) washing the membrane with water, to yield a hydrophilic membrane having
pores filled with a water-soluble gel sufficient to provide said
properties.
2. The method of claim 1 in which the membrane is a hollow fiber having a
lumen and the exterior of the membrane includes the lumen.
3. A module comprising a housing and a plurality of hollow fibers according
to claim 2.
4. In a method of carrying out affinity chromatography, the improvement
comprising employing hollow fibers according to claim 2.
5. The method of claim 4, in which, after step (h), a triazine dye is
pumped through the hollow fibers.
6. In a method of carrying out liquid-liquid extraction, the improvement
comprising employing hollow fibers according to claim 2.
7. In a method of carrying out gas separation using hollow fibers, the
improvement comprising employing hollow fibers according to claim 2.
8. The method of claim 2 in which the aqueous solution is pumped through
the lumens of the hollow fibers at a pumping rate between about 0.1 to
about 10 milliliters per minute.
9. The method of claim 2 in which the aqueous solution is removed from the
lumens of the hollow fiber membrane by pumping water through the lumens of
the hollow fiber at a pumping rate between about 0.1 to about 1 milliliter
per minute.
10. The method of claim 2 in which the crosslinking is catalyzed by
treatment with a base, and in which the base is pumped through the lumens
of the hollow fibers at a pumping rate between about 0.1 to about 10
milliliters per minute.
11. The method of claim 1 in which the polyol is polyvinyl alcohol.
12. The method of claim 1 in which the aqueous solution comprises polyvinyl
alcohol and divinyl sulfone.
13. The method of claim 12 in which the average molecular weight of the
polyvinyl alcohol is in the range of from about 5,000 to about 100,000
Daltons.
14. The method of claim 12 in which the polyvinyl alcohol is at least about
85% hydrolyzed from polyvinyl acetate.
15. The method of claim 12 in which the polyvinyl alcohol constitutes about
5 to about 15 percent of the aqueous solution and the divinyl sulfone
constitutes about 1 to about 5 percent of the aqueous solution, by weight.
16. The method of claim 1 in which the crosslinking is catalyzed by
treatment with a base.
17. The method of claim 1 in which the crosslinking is catalyzed by
treatment with heat.
18. A membrane produced by the method of claim 1.
Description
BRIEF DESCRIPTION OF THE DRAWINGS
Reference will be made to the accompanying drawings in which numerals
designate elements, and in which:
FIG. 1 illustrates the structure of the PVA/DVS gel.
FIG. 2 illustrates the apparatus used in Example 2 to assess the
performance of the gel-filled microporous membrane modules.
FIG. 3 illustrates resolution of a mixed solution with a gel stationary
phase hollow fiber membrane.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
Although any suitable microporous hollow fiber may be employed in the
practice of this invention, it is presently preferred to use microporous,
normally hydrophobic polyolefin (e.g., polypropylene or polyethylene;
polypropylene is most preferred) hollow fibers arranged generally parallel
to one another in a closely packed relationship within an outer shell
structure.
The hollow fibers employed in this invention may be, for example, those of
the type made using the "up-spinning" technique disclosed in U.S. Pat.
Nos. 4,405,688 and 4,541,981, each in the name of James J. Lowery et al,
and each being expressly incorporated herein by reference. Briefly,
non-porous precursor hollow fibers are produced according to the
techniques disclosed in these prior patents by melt spinning the precursor
fibers in a substantially vertically upward direction (i.e., up-spinning).
The thus melt spun precursor hollow fibers are then spin-oriented while
subjecting them to a symmetrical quenching step using a hollow annular
structure surrounding the precursor fiber which has one or more openings
on its inner surface that distribute the quenching medium against the
precursor fiber in a substantially uniform manner. The thus formed
precursor hollow fiber may then be heat-annealed by, for example,
subjecting the non-porous precursor hollow fiber to a temperature of
between about 5 degrees centigrade to about 100 degrees centigrade for a
time period of at least a few seconds (e.g., from a few seconds up to
about 24 hours, preferably between about 30 minutes to about 2 hours).
The finished microporous hollow fibers will possess an average inner
diameter in the range of from about 5 to about 1500 microns, and
preferably in the range of from about 70 to about 1500 microns. The fibers
are, moreover, characterized by a substantially uniform internal diameter
(I.D.), for example, a coefficient of variation in inner diameter through
a cross-section taken perpendicular to the axis of the fiber of less than
about 8%, preferably less than about 5%, and more preferably less than
about 3%.
The pores of the preferred microporous hollow fibers are essentially
interconnected through tortuous paths which may extend from one exterior
surface or surface region to another, i.e., open-celled. Further, the
pores of the preferred microporous hollow fibers of the invention are
microscopic, i.e., the details of the pore configuration are described
only in terms of microscopic dimensions. Thus, the open cells or pores in
the fibers are smaller than those which can be measured using an ordinary
light microscope, because the wavelength of visible light, which is about
5,000 Angstroms, is longer than the longest planar or surface dimension of
the open cell or pore. The pore size of the microporous hollow fibers may
be defined by using electron microscopy techniques which are capable of
resolving details of pore structure below 5,000 Angstroms or by mercury
porosimetry techniques.
The average effective pore size of the microporous hollow fibers useable in
the practice of this invention is preferably between about 50 to 2,000
Angstroms, and more typically between 100 to 1,000 Angstroms. By
"effective pore size" is meant the smallest dimension of a pore which
would allow a generally spherical particle of that same dimension to pass
therethrough. The pores generally have an elongated shape with a width of
from about 50 to 2,000 Angstroms, and a length of from about 500 to 10,000
Angstroms. Hence, the "average effective pore size" of the preferred
microporous hollow fibers will usually be determined by the width
dimension of the pores. These pores will, moreover, be fairly uniform
around the circumference of the fiber. For example, the preferred
microporous hollow fibers will exhibit an average ratio of the maximum
pore density to the minimum pore density around the circumference of the
fiber of less than about 3:1, and usually less than about 2:1.
Three further criteria for the preferred hollow fibers to be used in
chromatographic applications are as follows (1) the fiber wall thickness
is preferably less than 20% of the fiber diameter, more preferably less
than 10% of the fiber diameter; (2) diffusion in the stationary phase
should be reduced no more than a factor of five from the mobile phase; and
(3) the diameters of the fibers should be consistent to within 5%
variability. It should be recognized, however, that other hollow fibers
can be used.
Microporous hollow fibers of the type described above are commercially
available from Hoechst Celanese Corporation, Separations Products
Division, Charlotte, N.C., under the registered trademark CELGARD.
The microporous hollow fibers are preferably a part of a module having an
inlet end and an outlet end. The module includes a number (e.g., from a
few hundred to many thousand) hollow fibers of predetermined length
arranged substantially parallel to one another within the central space of
a generally tubular shell structure (e.g., glass, metal, or plastic
tubes). The individual fibers are positionally retained within the outer
shell structure by means of suitable potting compounds (e.g., epoxy
resins). A particularly preferred epoxy resin is a 2.5:1.0 mixture (by
volume) of FE-5045A and FE-5045B, both commercially available from H.B.
Fuller, St. Paul, Minn. Modules of this type are commercially available in
a wide range of sizes and capacities. For example, one particular
microporous hollow fiber module which may be satisfactorily employed in
the practice of the present invention is commercially available from the
Hoechst Celanese Corporation, Separations Products Division, Catalog No.
50101060. This module has 27,000 CELGARD(R) microporous hollow fibers of
100 micron internal diameter. Other suitable microporous hollow fiber
modules may, however, be used for the purposes of this invention.
In preferred embodiments of this invention, hollow fibers are supported in
glass tubes ranging from 20 centimeters to 60 centimeters in length,
having an outside diameter of 6.0 millimeters and a glass wall thickness
of 1.0 millimeter.
A key factor in designing a module for use according to this invention is
the expected throughput. The throughput is directly proportional to the
lumen volume (and hence, the number and length of fibers), but as the
cross-sectional area of the module increases, it becomes increasingly
difficult to evenly distribute the mobile phase and sample among the
hollow fibers. As the column length increases, the capacity to separate
the mobile phase into its component parts increases but at the expense of
increased pressure drop. According to preferred embodiments, the column
length is kept at a reasonable length and the cross-sectional area
relatively large, as the best compromise of these tradeoffs. In this
manner, the pressure drop is manageable, and the column does not become
cumbersome. In a particularly preferred embodiment, the column length is
about 5 meters, and the operating pressure drop is one pound per square
inch.
The coating process is generally applicable to any configuration of
microporous hollow fibers. Microporous hollow fibers having an inner
diameter of both 240 and 100 microns and fiber wall thickness of about 30
microns, for example, have been successfully used. However, for reasons
not completely understood, 240 micron hollow fibers give more consistently
uniform coatings. Applicants believe that the fiber porosity (40% for the
240 micron fibers and 30% for the 100 micron fibers), and/or the smaller
internal diameter of the 100 micron fibers (possibly leading to lumen
clogging by coating polymer) are the likely reasons why the larger
diameter hollow fibers give better results.
Microporous hollow fibers having nominal inside diameters of 240 microns
and 100 microns are respectively available under the tradenames CELGARD(R)
X-20 and CELGARD(R) X-10 from the Hoechst Celanese Corporation, Charlotte,
N.C. These fibers have a wall thickness of about 30 microns.
Each fiber is a small tube with a microporous wall. The porosity of the 100
micron and 240 micron inside diameter fibers are 30% and 40%,
respectively. The pores are small rectangular slits with channels
extending through the fiber walls.
Once suitable microporous hollow fibers (MHF) have been selected, a module
should be constructed incorporating them.
In a preferred embodiment, the glass modules are provided with two 6 mm
ports each about 2 cm in length, at a point about 3 cm from each end of
the module. These ports permit application of a driving force of up to
about one pound per square inch, (preferably about one-half pound) across
the membrane, which aids fiber wetting. It is not essential that modules
having ports at the ends be used. A simple glass tube is often easier to
work with. However, it may be difficult to completely fill the shell side
with a polymer solution without ports.
Having completed preparation of the module, one end is then placed in a
standard 1/4" chromatographic fitting, which is then connected to a
suitable peristaltic pump. The coating procedure described below is then
carried out by pumping via the fitting.
The first step in the coating process is to make the hydrophobic membrane
water-wettable. This is done by treating the fiber with a surfactant
solution.
Suitable surfactants are well known to those skilled in the art, and any
surfactant typically used to wet hydrophobic microporous membranes can be
used. Particularly preferred is a solution of TWEEN(R) 60, water and
methanol.
The surfactant solution is then pumped into the module and through the
lumens to wet the fibers. The pumping rate should be moderate, say about
one milliliter per minute (ml/min) and preferably about 3ml/min. The
pumping is continued until the solution completely wets the fibers and
fills both ports.
Next, the module is prepared for the polymer coating process. The
surfactant solution is drained from the shell side, and the module is
dried. Drying can be carried out in any suitable manner, preferably by
either air drying for say, 15-20 hours or by placing the module in a
vacuum oven.
The polymer solution consists of polyvinyl alcohol (PVA), divinyl sulfone
(DVS) and water. The solution is mostly distilled water, because a dilute
solution is essential in order to facilitate rapid diffusion into the
lumen pores. The use of distilled water rather than, e.g. tap water, is
preferred but not essential. Preferably, the water constitutes at least
about 85% of the solution, more preferably at least about 90%, and most
preferably about 93.3%.
Although the discussion herein is directed to PVA/DVS gel systems, it
should be understood that other polyol/DVS-type gellable systems can also
be used. Suitable polyol/DVS systems are disclosed, for example, in the
Handbook of Fiber Science and Technology: Volume II--Chemical Processing
of Fibers and Fabrics--Functional Finishes Part A (Ed. by Menachem Lewin
et al), (Marcel Dekker, N.Y. 1983), pp. 23-28, which is hereby
incorporated by reference. Of particular interest is the disclosure in
this reference of cellulosic materials treated with divinyl sulfone and
divinyl sulfone precursors. Most broadly, the coating compositions of this
invention are made up from at least one polyol, such as, but not limited
to, PVA or the polyols disclosed in the above-referenced Handbook,
combined with divinyl sulfone and/or one or more divinyl sulfone
precursors as also disclosed in the Handbook.
The PVA and DVS must be suitably selected to result in a gel to fill the
substrate membrane pores. Generally, the PVA should have a sufficiently
high average molecular weight to facilitate gelation; and low enough to
avoid clogging the lumens of the hollow fibers. For hollow fibers having
particularly small lumens, the PVA should have a relatively low average
molecular weight. Concurrently, as the molecular weight of the PVA
decreases, the concentration of DVS employed should be accordingly
increased. The resulting polymer should have a sufficiently high molecular
weight to facilitate entanglement of the polymer in the pores of the
substrate membrane, yet low enough so that the polymer can swell into a
gel. Those skilled in the art will appreciate that these parameters may
need to be adjusted in particular cases to result in an effective gel.
The PVA should generally be characterized by an average molecular weight of
at least about 5,000 and not more than about 100,000; preferably the PVA
has a molecular weight ranging from about 10,000 to about 100,000, and
most preferably from about 10,000-30,000 Daltons. PVA is often obtained by
hydrolyzing polyvinyl acetate. For the purposes of this invention, the PVA
should be at least 85% hydrolyzed, preferably at least 95%, and most
preferably at least 98%. A suitable product is AIRVOL 107, which is
commercially available from Air Products and Chemicals Inc.
The PVA should generally constitute about 5 to about 15%, preferably about
5 to about 10%, and most preferably about 5.6% of the solution.
Divinyl sulfone is a standard item of commerce. It should generally
constitute about 1 to about 5%, preferably about 1 to about 2%, and most
preferably about 1.1% of the solution.
The proportions of PVA and DVS should be adjusted to yield the appropriate
percentage of unreacted alcohol groups on the polymer. Preferably, about
90%, and most preferably about 93% of the alcohol groups should remain
unreacted.
The polymer solution is mixed and then pumped through the lumens of the
hollow fibers. The pumping rate should generally be moderate, say, about
0.1 to about 10ml per minute, preferably about 0.1 to about 1 ml per
minute. The polymer solution is pumped through the lumens until the fibers
are completely wetted and both ports are filled. At this point it is
convenient to cap the ports, e.g. with standard 1/4" SWAGELOK caps.
Clearing the excess polymer solution from the lumens is critically
important. The polymer must be removed from the lumens without washing it
out of the lumen pores. This is accomplished by passing distilled water
through the module at a low flow rate, generally about 0.1 to about 1 ml
per minute, preferably about 0.5ml per minute, until the viscosity of the
elutant decreases. This change in viscosity can be readily observed by the
naked eye because the polymer solution is several times more viscous than
water, and therefore flows more slowly. Also, the polymer solution is
light yellow in color. The displacement of the excess polymer generally
takes about 2-4 minutes. Typically a volume of water equal to about half
the volume of the lumen should be employed.
Crosslinking of the PVA/DVS is then achieved by introduction of a catalyst.
A basic solution is passed through the lumens at a moderately heated
temperature. Preferably, sodium hydroxide (NaOH) is employed. The base
should generally be heated to about 40-60 degrees centigrade, preferably
about 50 to 60 degrees, and most preferably to about 55 degrees
centigrade. A particularly preferred solution consists of 80% distilled
water and 20% NaOH. As will be readily understood by those skilled in the
art, other catalyst systems may be used, and heating to aid the
crosslinking reaction may not be necessary. In particular, systems
employing DVS as the crosslinking agent may not require heating and/or a
catalyst.
The catalyst solution is pumped into the fiber lumens, at a low flow rate,
generally about 0.1 to 10 ml per minute, preferably about 0.1 to 1 ml per
minute, and most preferably about 0.5 ml per minute. The gel begins to
form within seconds after applying the base, turning the fibers a light
yellow. The base is circulated for several minutes, generally about 3-4
minutes. The gelation occurs by a Michael's addition reaction (see FIG.
1).
The module is then washed with distilled water. For the modules described
here, about 2 liters is sufficient. The flow rate should be moderate, say
about 1 to about 10 ml per minute, preferably about 2 ml per minute. The
fibers lose their yellow color soon after the washing begins. In order to
completely clear the lumen, it is helpful to run the washing at a high
flow rate near the end, say, about 8 ml per minute.
The resulting module is characterized by pores which are filled with a
PVA/DVS aqueous gel. The structure of the gel is shown in FIG. 1.
In use, the hollow fiber modules of the invention are first charged with a
solution to be separated. The solution is generally charged under
pressure, in order to maximize the transport rate of the solution through
the lumens of the hollow fibers.
The modules can be used either singly or in parallel. Parallel operation
permits increased total throughput without the problem of increased
pressure drop.
The pore-filled microporous hollow fiber membrane modules of the invention
can be used in a broad range of chromatographic separations applications,
including both differential migration for small molecules (e.g.,
liquid-liquid extractions), and affinity adsorption for charged species,
particularly proteins and enzymes. The pore-filled hollow fibers
constitute stabilized, immobilized liquid membranes having utility
generally where such membranes are needed. The stability of the
immobilized membranes permit operation at high pressures without resultant
bleed-out of the immobilized liquid membrane, yet without reduced flow
rates, because diffusion migration of the mobile phase is not hindered.
While the invention has been described only in connection with hollow
fibers, the pore-filling process could also be applied to other forms of
microporous membranes, particularly flat sheet membranes.
EXAMPLE 1
Microporous hollow fibers (CELGARD(R) X-20) were potted in glass tubes
using an epoxy resin. Two tubes, 20 cm and 60 cm in length, respectively,
were employed. The tubes each had an outer diameter of 1/4", and included
two ports having outer diameters of 60 mm placed approximately 11/4" from
each end. After the epoxy was cured, one end of each glass module was
placed in a standard 1/4" chromatographic fitting, in this case a Valco
fitting. The fitting was connected to a peristaltic pump.
A solution was prepared consisting of one gram of TWEEN(R) 60, a
commercially-available polyoxyethylenesorbitan, 40ml distilled water, and
60ml of methanol. The solution was pumped into the modules and through the
lumens of the fibers at a rate of about 3ml per minute, wetting the fibers
almost immediately. The pumping was continued until the solution
completely wet the fibers and filled both ports. The solution was then
drained from the shell side, and air was passed through the module for
15-20 hours to dry the module.
Next, a polymer solution (100 ml) was prepared consisting of 93.3%
distilled water, 5.6% polyvinyl alcohol (Airvol 107, average molecular
weight 10,000-30,000; 98% hydrolyzed from polyvinyl acetate) and 1.1%
divinyl sulfone (all percentages by weight). This solution was then pumped
through the fiber lumens until the fibers were completely wetted and both
ports were full. Caps (1/4" Valco) were placed on the ports and hand
tightened.
Distilled water was then pumped through the modules at a flow rate of about
0.5ml per minute until the polymer solution was displaced from the lumens.
This took about 2-4 minutes.
A solution consisting of 20% NaOH and 80% distilled water (by weight) was
prepared (100 ml), heated to 55 degrees centigrade and pumped through the
fiber lumens at a rate of about 0.5 ml per minute. The fibers turned a
light yellow, indicating the formation of a gel. Pumping of the base
continued for 3-4 minutes.
The modules were then washed by passing about 2 liters of distilled water
through the lumens at about 2.0 ml per minute. The fibers lost the yellow
color after about 20 ml of this washing. In order to completely clear the
lumens, the washing was run at a high flow rate (about 8 ml per minute)
for the last hour of washing.
EXAMPLE 2
An experimental system used to evaluate the performance of the modules
prepared in Example 1 is schematically shown in FIG. 2. An aqueous
solution of several solutes is prepared in parts by weight. The amount of
total solutes to be injected into the module is kept well below overload
conditions, by preparing a dilute solution and by using injection volumes
of 10-20 microliters. An injection valve is primed with sample solution
and the solution to be separated is then pumped into the module. The
solution (mobile phase) travels through the hollow fiber lumens and into a
UV-visible light detector, which sends a voltage signal to an integrator.
The detector normally uses wavelength settings of 254 nanometers for all
solutes except ketones, which are measured at 265 nanometers. The
resulting data are equivalent to concentration readings. The integrator
stores the signal as a function of elapsed time. In these trials, a
SpectraPhysics Isochrom pump, a Scientific Systems Inc. pulse dampener
(model LP-21), a Milton-Roy Variable Wavelength Detector (model 3010), a
Rheodyne 7010 injection valve, and a Hewlett-Packard Integrator (model
3396A) were employed. This equipment is all standard; other equipment
could readily be substituted.
A module 50 cm in length, containing 120 fibers having inside diameters of
240 microns and a fiber wall thickness of 25.4 microns was prepared
according to Example 1 and used to separate an aqueous mixture of acetone,
phenol and m-nitrophenol. The solution to be separated was passed through
the fibers at a flow rate of 0.00083 cubic centimeters per second. The
results are plotted in FIG. 3. As can be seen, the m-nitrophenol was
completely separated from the other solution components, and the other two
mixture components were substantially separated from each other.
EXAMPLE 3
Additional modules were prepared according to Example 1 and tested in the
same manner described in Example 2. The results are shown in TABLE 1.
In affinity adsorption mode, separations of proteins and enzymes may be
carried out by the selective binding of ligands for the respective
proteins and enzymes, in the immobilized gel. The PVA gel has hydroxy
groups which are available for derivatization. The ligands to be bound
should preferably be inexpensive, because large amounts will be needed if
preparative chromatography is to be carried out in addition to analytical
chromatography (the former refers to preparation of useable quantities of
a desired product).
A preferred group of ligands in the case of proteins are the triazine dyes.
These are referred to as pseudo-ligands, since they are not natural
protein-binding substances, but rather mimic the same. Triazine dyes can
be attached to the PVA gel by forming an ether linkage via a hydroxy
group. These dyes are suitable for large-scale affinity separations for
several reasons. These dyes have a greater capacity for binding proteins
than naturally-occuring ligands; they can bind anywhere from 10 to 100
times more than a true affinity ligand. The natural ligands are vastly
more expensive than the dyes, which are available as commodity chemicals
in large quantities and at low costs. Most importantly, however, the dyes
have a propensity to bind, selectively and reversibly, a plethora of
proteins. There are two kinds of triazine dyes, the Procion MX and Procion
H groups. Although either can be used, the Procion MX dyes are generally
more reactive with the hydroxy groups of the gel, making application of
heat less necessary and possibly avoidable in binding the dye to the gel.
The triazine dyes are particularly effective in the purification of
pyridine nucleotide-dependent dehydrogenases, kinases, coenzyme
A-dependent enzymes, hydrolases, acetyl-, phosphoribosyl-, and
amino-transferases, RNA and DNA nucleases, decarboxylases,
sulfohydrolases, phosphorylase, myosin, serum albumin, clotting factors,
lipoproteins, complement proteins, and interferon.
The following Example demonstrates suitable procedures for binding a
triazine dye psuedo-ligand onto a PVA gel immobilized in hollow fibers.
EXAMPLE 4
Hollow fibers having a PVA/DVS gel immobilized in the pores were prepared
as in Example 1. A solution consisting of 0.6 grams of Reactive Blue 4
(Sigma Chemicals, R-9003) containing approximately 40% of the Procion MX-R
dye was dissolved in 40 ml of distilled water and 20 ml of a 4 molar
sodium hydroxide solution. After dissolution of the dye, 0.5 ml of an
aqueous 20% by weight sodium hydroxide solution was added to the mixture
This solution was immediately pumped into the module in order to avoid
hydrolysis of the chloro groups by the base. The module employed was 60 cm
in length containing 132 hollow fibers with nominal inside diameters of
240 microns. This module had a lumen volume of 3.6 cubic centimeters. The
solution was pumped through the module for 15 minutes at a rate of 0.3 ml
per minute. Flow was halted, and the module was capped at the ends. After
the module sat for two hours, distilled water was passed through the
module until the module effluent was no longer blue (usually about 1
liter). The above procedure was repeated in order to ensure adequate dye
coverage on the gel. The fibers were then washed copiously and
sequentially with (1) water; (2) 1 molar sodium hydroxide/25% ethanol; (3)
water; (4) 1 molar sodium hydroxide/0.2 molar phosphate, pH 7.0; (5)
water. Inspection of the module showed a uniform blue color in the hollow
fibers. At this point the module was ready to perform affinity
separations.
In using a module prepared as above for affinity chromatographic
separations, four distinct steps are involved. First, the solution to be
separated is injected into a noneluting buffer and carried into the hollow
fibers The solutes in the solution that "recognize" the affinity ligand
are bound to the gel. Second, inert or weakly-retained species are removed
by passing a noneluting buffer through the hollow fibers. Third, the
elution is carried out by changing the composition of the mobile phase.
While the binding of the solute to the ligand is reversible, the elution
can only occur by changing some property of the mobile phase. It is
convenient to install a valve upstream of the hollow fiber module which
will allow selection of several different mobile phase compositions.
Fourth, the module must then be returned to its initial condition before
another solution is injected.
Five ways of changing the mobile phase are as follows: changing the pH,
increasing the ionic strength, adding a chaotropic agent, adding organic
solvents, or adding a competing biologically specific ligand. Sometimes
combinations of these changes are necessary to elute the solute. The most
selective method is the addition of competing ligands.
The following Example shows the utility of a typical module for both
differential adsorption and affinity chromatography:
EXAMPLE 5
A module is prepared containing 82,500 hollow fibers having pores filled
with PVA/DVS gel prepared and deposited as in Example 1. In this module,
the cross-sectional area will be about 100 square centimeters (or a column
diameter of about 10 centimeters). This module design will allow for the
isolation of two low molecular weight solutes (molecular weight less than
500 ), at a throughput of approximately 37 grams per day for each solute
where the selectivity of the separation arises from differential
absorption/desorption and the differences in partitioning between the
solutes is a factor of 2. The same device, with an immobilized triazine
dye in the gel, isolates approximately 175 grams of bovine serum albumin
per day when operated in the affinity mode.
The use of hollow fibers to carry out liquid-liquid extractions is
disclosed in Sirkar U.S. Pat. No. 4,789,468, the entirety of which is
hereby incorporated by reference. That patent discloses the application of
static pressure to maintain the interface between the two liquid phases at
the membrane. Such procedures can be inconvenient, especially in large
modules where the pressure drop in the module may be as big as the static
pressure difference. Additionally, surface active species may, despite the
static pressure, enter and "wet out" the membrane, resulting in a
compromised extraction. By using the hollow fibers of this invention
having gel-filled pores, these problems are avoided because the gel
prevents either the feed or extractant liquid/phases from entering the
hollow fiber substrate membranes.
The gel-filled hollow fiber membranes can also be used to carry out gas
separations. In contrast, for example, to glassy polymers such as modified
or unmodified polycarbonates or polyimides, the gelled polymers of the
invention provide a matrix having a higher diffusion rate, resulting in
more rapid and higher volume gas separations. Particular gel-forming
materials within the scope of the above discussion may be selected by
those skilled in the art for particular separations applications.
While the invention has been described in connection with what are
presently considered to be the most practical and preferred embodiments,
it is to be understood that the invention is not limited to the preferred
embodiments, but rather is intended to cover various modifications and
equivalents included within the spirit and scope of the appended claims.
TABLE 1
______________________________________
RESULTS WITH PVA GEL STATIONARY PHASE
MODULE.sup.1
SOLUTE A.sup.2 B.sup.3
C.sup.4
D.sup.5
______________________________________
s s s.sup.2
196/100/37.0
phenol 690 4,220 420,000
5.11
1,380 8,310 1,060,000
5.03
acetone 690 1,932 118,000
1.80
1,380 4,000 371,000
1.90
120/240/51.5
phenol 696 1,956 96,800 1.82
3,468 10,020
745,000
1.89
m-nitrophenol
696 4,950 648,000
6.13
3,468 24,840
4,640,000
6.16
acetone 696 1,278 34,560 0.85
3,468 6,612 252,000
0.91
2-pentanone
696 1,346 48,600 0.95
3,468 6,954 337,000
1.00
blue dextran
696 702 14,200 0.008
3,468 3,480 130,000
0.003
132/240/62.0
phenol 4,450 11,580
1,010,000
1.60
1,482 3,780 223,500
1.55
m-nitrophenol
4,450 23,900
5,600,000
4.37
1,482 7,740 2,100,000
4.22
acetone 4,450 8,000 363,000
0.80
1,482 2,650 115,000
0.79
120/240/21.5
phenol 281 714 33,200 1.55
m-nitrophenol
281 1,550 184,000
4.51
acetone 281 498 10,800 0.77
______________________________________
.sup.1 number of fibers/nominal fiber i.d. in microns/module length in cm
.sup.2 mean residence time of the mobile phase
.sup.3 average solute residence time
.sup.4 column variance
.sup.5 capacity factor
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